This project proposes to isolate and characterize the subcellular structures of vascular smooth muscle responsible for the determination of the intracellular calcium level (and hence the amount of tension development). In particular we propose to measure the following properties of four distinct subcellular organelles obtained by extraction and differential centrifugation of bovine aortic muscularis: Sarcoplasmic Reticulum (SR) - the capacity and affinity of the active calcium uptake mechanism associated with the isolated SR vesicles. Also the associated Ca-ATPase will be measured. Mitochondria - the capacity and affinity of the active calcium uptake mechanism associated with the mitochondria. Myofibrils - the Ca and Mg-ATPase activity as a function of both calcium and magnesium concentrations. Also the calcium binding will be measured and attempts to determine the precise nature and location of the calcium binding site have been initiated. Sarcolemma -Attempts to isolate this organelle in vesicular form will be performed. If successful, the calcium affinity and capacity will be measured. The resultant data from these studies will be incorporated into a systems description for the regulation of intracellular calcium. To check this systems description, we will measure the effects of various vasoactive agents such as norepinephrine and procaine on the four elements of the system and see if these effects would cause a change in the intracellular calcium consistent with the appropriate contractile response to the agent. BIBLIOGRAPHIC REFERENCES: Ford, George D. and Hess, Michael L. Calcium accumulating properties of subcellular fractions of vascular smooth muscle. Circ. Res. 37: 580-587 (1975). Soulsby, M E., Hess, M.L. and Ford, G.D. The effect of endotoxin on calcium uptake by mitochondria from cardiac and vascular smooth muscle. Fed. Proc. 34:380 (1975). (Abstract).